HDL levels are inversely related to coronary artery disease. The HDL receptor, SR-BI, plays a key role in HDL metabolism. SR-BI binds HDL and mediates the process known as selective lipid uptake, the mechanism of which is not understood. Various studies have indicated that selective uptake by SR-BI in certain cells occurs at the cell surface by a non-endocytic process. However, recent studies have reported that in hepatocytes SR-BI mediates the internalization of intact HDL particles and suggest that selective lipid uptake involves HDL internalization and recycling in cells. We have shown that SR-BII, a variant of SR-BI, differs markedly from SR-BI in its cellular trafficking and endocytosis of HDL. The central hypothesis of this proposal is that the functions of SR-BI and SR-BII in intracellular cholesterol trafficking are coupled to receptor/ligand recycling in cells and that SR-BI and SR-BII follow different internalization and recycling routes and differentially affect cellular cholesterol trafficking and regulation. The specific aims are: 1) determine the contribution of HDL internalization and intracellular recycling to the selective lipid uptake process mediated by SR-BI and SR-BII. HDL recycling by each of the two receptors will be quantified in hepatocytes and other cells, 2) elucidate the mechanisms responsible for HDL internalization and recycling by SR-BI and SR-BII. Endocytic and recycling pathway(s) used by SR-BI and SR-BII will be studied and protein motifs in the C-terminal tails of SR-BI and SR-BII that are responsible for receptor targeting will be identified, 3) determine how differences in the intracellular targeting of HDL by SR-BI and SR-BII influence cellular cholesterol regulation and trafficking. We will determine the effects of selective lipid uptake via SR-BI and SR-BII on the pool of regulatory sterols in cells and on cholesterol efflux and 4) assess the roles of SR-BI and SR-BII in HDL metabolism and atherogenesis in vivo using isoform-specific knock-in mice. Isoform-specific knock-in mice will be generated and the effects of each receptor on plasma lipoproteins, HDL metabolism and atherogenesis studied. [unreadable] [unreadable]